Replies

• 1

  Bronwen Dekker

  15 August 2008 | 15:52

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  We have a protocol in the peer-reviewed section of our content that might be of use to you:

  Purification of antibodies using the synthetic affinity ligand absorbent MAbsorbent A2P

  From the abstract it might be specific to antibodies from ovine serum, but I cannot think at the moment why it could not be adapted to others.

  Parts of the protocol that you don’t need a site-licence to access:

  Table 1. MAbsorbent A2P—product specifications

  Table 2. Overview of purification steps

  Table 3. Troubleshooting table

  Figure 1

  Figure 2

• 2

  Lindy Abas

  06 November 2008 | 22:09

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  If you can run some of the recombinant protein on a gel (use a preparatory comb or use the whole width of the gel with no comb at all), then blot the gel, stain the membrane using Ponceau, identify the band and cut it out from the blotting membrane. Use this cut-out piece to affinity purify your specific antibodies, see Smith & Fisher (1984) J. Cell Biol. 99: 20-28. You will need about 100 micrograms of the protein. If you can’t get the recombinant protein pure or highly expressed enough to identify it as a distinct band, then why not use the opposite approach, i.e. remove all the antibodies against the non-specific E.coli proteins. Try blotting e.coli protein extract (i.e. with NO recombinant protein expression) onto a membrane as above, then use the whole membrane to preabsorb your serum. Sorry this is a late answer!
  lindy